DESCRIPTION: This is a proposal to investigate the biochemical mechanisms of cell necrosis. The mitochondrial permeability transition (MPT) has been established as a key event in hepatocyte cell death caused by agents that inhibit mitochondrial electron transport and in cell death in L929 fibroblasts caused by tumor necrosis factor (TNF). The objectives of the proposal are to identify events leading to the MPT and to determine how induction of the MPT ultimately causes changes in the cytoskeleton and the plasma membrane that result in necrosis. The first specific aim is to establish the mechanism of induction of the MPT. The role of long-chain fatty acyl CoA in de-energizing mitochondria will be investigated in hepatocytes, whereas the roles of mitogen activated protein kinases (MAPKs) will be investigated in the L929 cells. Specific aim 2 will investigate the hypothesized coupling of the MPT to changes in cytoskeletal proteins. Both biochemical and morphologic studies are proposed. In Specific aim 3, the role of phospholipase activation in causing plasma membrane alterations will be investigated. These studies will employ phospholipase A2 inhibitors and analyses of phospholipase activities in plasma membrane fractions. In Specific aim 4, changes in the molecular order of plasma membranes will be investigated with spin labeling methods and changes in ion permeability will be examined.